Peripheral arterial disease (PAD) and its severe form critical limb ischemia (CLI) affect nearly 8 million Americans. Most patients with chronic metabolic and cardiovascular diseases such as diabetes also suffer from PAD/CLI. Patients with PAD/CLI suffer from debilitating symptoms such as immobility, ulceration, impaired wound healing, and intractable ischemia. The pathology of PAD/CLI involves vascular regression in the skeletal muscle, which leads to significant myopathy. Treatment is limited to surgical reconstruction, which is often costly and ineffective leading to limb amputations. 'Therapeutic angiogenesis' to improve muscle vascular supply is an urgent area of research in PAD/CLI. Within this field, focus has been on pro-angiogenic regulators such as vascular endothelial growth factor-alpha (Vegfa) and hypoxia inducible factor 1 (HIF1), and how they can improve muscle vasculature. Unfortunately, individual angiogenic factors and even HIF1 have so far been ineffective targets in PAD/CLI. Notably, skeletal muscle vascular supply and neo angiogenesis is controlled in a paracrine fashion by both pro-angiogenic and anti-angiogenic factors secreted by the muscle. While molecular regulation of pro-angiogenic factors such as Vegfa in the muscle is defined, surprisingly, the molecular regulation of anti-angiogenic factors is poorly defined. We have preliminarily found that a transcriptional cofactor peroxisome proliferator-activated receptor gamma co-activator 1 beta (PGC1?), which typically functions by activating nuclear hormone receptors, encodes an inhibitory angiogenesis gene program in the skeletal muscles. This program constitutes induction of anti-angiogenic factors and repression of proangiogenic factors, resulting in a net inhibition of skeletal muscle revascularization in ischemia. Interestingly, we found that PGC1? expression is induced along with the anti-angiogenic factors in diabetic muscles, which are characterized by the pathological symptoms of PAD/CLI such as vascular regression, ischemia and impaired neo-angiogenesis. Therefore, the first objective of this research proposal is to decipher the role of endogenous muscle PGC1? in regulating inhibitory angiogenic program and its effect on ischemic muscle revascularization and diabetic muscle angiopathy. The second objective is to establish COUP-TFI (chicken ovalbumin upstream promoter transcription factor I) as the nuclear receptor involved in PGC1?-driven inhibitory angiogenesis. Our preliminary data indicates that the anti-angiogenic effects of PGC1? may be mediated by activating nuclear receptor COUP-TFI. Our central hypothesis is that PGC1? in the skeletal muscle instigates a paracrine anti-angiogenic gene program by activating COUP-TFI, blocks ischemic neo angiogenesis, and contributes to diabetic muscle angiopathy. The specific aims to test this hypothesis are to: (Aim 1) Investigate the role of endogenous PGC1? in muscle angiogenesis and (Aim 2) Investigate whether PGC1? and COUPTFI interact to regulate muscle angiogenesis. In Aim 1, we will use muscle-specific deletion of PGC1? in mice to investigate how the loss of muscle PGC1? affects muscle vasculature, ischemic revascularization and the expression of angiogenic factors. In Aim 2, we will first perform in vitro experiments to study the interaction between PGC1? and COUP-TFI, and how they regulate muscle angiogenic genes. Second, we will use muscle-specific compound transgenic mice to investigate whether the anti-angiogenic effects of musclespecific PGC1? over-expression is lost on muscle-specific COUP-TFI deletion. We will also investigate the impact of double muscle-specific PGC1? and COUP-TFI knockout on muscle neo-angiogenesis and revascularization. The significance of our work is that it aims to discover how anti-angiogenic gene program is regulated in the skeletal muscle, and how it affects revascularization in muscle ischemia and diabetic muscle angiopathy. Our research is innovative as we will study the unexplored area of anti-angiogenic gene programming in the treatment of PAD/CLI. The findings will have implications for devising new treatment for muscle ischemia in PAD/CLI, and broadly in other diseases such as cardiac infarction, stroke and tumor angiogenesis.